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Genetic Mutations (Review)
Recall: The Central Dogma
Normal gene
DNA (antisense strand)
mRNA
Polypeptide
GGTCTCCTCACGCCA
↓
CCAGAGGAGUGCGGU
Codons
↓
Pro-Glu-Glu-Cys-Gly
Amino acids
The antisense strand is the DNA strand which acts as
the template for mRNA transcription
What Causes Mutations?
• There are two ways in which DNA can
become mutated:
– Mutations can be inherited
• Parent to child (germline mutation)
– Mutations can be acquired
• Environmental damage
• Mistakes when DNA is copied (somatic
mutation)
Significance of Mutations
• Most are neutral
• Eye color
• Birth marks
• Some are harmful
• Sickle Cell Anemia
• Down Syndrome
• Some are beneficial
• Sickle Cell Anemia to Malaria
• Immunity to HIV
• Point Mutations – change in one
nucleotide
– Substitution
• THE FAT CAT ATE THE RAT
• THE FAT HAT ATE THE RAT
– Insertion (Frameshift)
• THE FAT CAT ATE THE RAT
• THE FAT CAT HATE THE RAT
– Deletion (Frameshift)
• THE FAT CAT ATE THE RAT
• THE FAT CAT AT THE RAT
__________________________________________________
Substitution
Normal gene
GGTCTCCTCACGCCA
↓
CCAGAGGAGUGCGGU
Codons
↓
Pro-Glu-Glu-Cys-Gly
Amino acids
Substitution mutation
GGTCACCTCACGCCA
↓
CCAGUGGAGUGCGGU
↓
Pro-Arg-Glu-Cys-Gly
Substitutions will only affect a single codon
Their effects may not be serious unless they affect an
amino acid that is essential for the structure and function of
the finished protein molecule (e.g. sickle cell anaemia)
Nonsense
Normal gene
GGTCTCCTCACGCCA
↓
CCAGAGGAGUGCGGU
Codons
↓
Pro-Glu-Glu-Cys-Gly
Amino acids
Substitution mutation
GGTCTCCTCACTCCA
↓
CCAGAAGAGUGAGGU
↓
Pro-Glu-Glu-STOP
_________________________________________________
No change
Normal gene
GGTCTCCTCACGCCA
↓
CCAGAGGAGUGCGGU
Codons
↓
Pro-Glu-Glu-Cys-Gly
Amino acids
Substitution mutation
GGTCTTCTCACGCCA
↓
CCAGAAGAGUGCGGU
↓
Pro-Glu-Glu-Cys-Gly
The genetic code is degenerate
(What does this mean?)
Degenerate = ________________________________
Changes in the third base of a codon often have no
effect.
• Frameshift Mutations – shifts the
reading frame of the genetic
message so that the protein may
not be able to perform its
function.
– Insertion
• THE FAT CAT ATE THE RAT
• THE FAT HCA TAT ETH ERA T
H
– Deletion
• THE FAT CAT ATE THE RAT
• TEF ATC ATA TET GER AT
H
Inversion
Inversion mutations, also, only affect a small part of the
gene
Normal gene
GGTCTCCTCACGCCA
↓
CCAGAGGAGUGCGGU
Codons
↓
Pro-Glu-Glu-Cys-Gly
Amino acids
Inversion mutation
GGTCCTCTCACGCCA
↓
CCAGGAGAGUGCGGU
↓
Pro-Gly-Glu-Cys-Gly
What happens to “bad” mutations overtime?
Why can’t we get rid of “bad” mutations?
What happens to “good” mutations?
Mutation Example 1:
Sickle Cell Anemia
Sickle cell anemia
Image Credit: http://explore.ecb.org/
Blood smear (normal)
Image Credit: http://lifesci.rutgers.edu/~babiarz/
Mutations of haemoglobin
• Haemoglobin is a tetramer = 2  and 2 -chains
• The genes for these polypeptides are found on
different chromosomes
• The -chain gene is found on chromosome 11
• The -chain gene is found on chromosome 16
• Several inherited diseases occur on the -chain,
which contains 146 amino acids.
Mutation
S (sickle cell
anaemia)
Codon Change to DNA
sense strand
6
GAG to GTG
Change in
Amino Acid
Glu to Val
Why does sickle cell anemia persist in
the population?
• The malarial parasites grow poorly in red
blood cells from either homozygous sickle-cell
patients or healthy heterozygous carriers
• Malaria is rarely found among carriers of this
mutation
• Malaria has served to maintain the otherwise
deleterious sickle-cell mutation at high
frequency in regions of Africa
Mutation Example 2:
Cancer Genes
• Cancer genes are causally implicated in
oncogenesis
• Mutations in cancer genes can occur somatically
or can be inherited
• Somatic mutations can occur in any of the cells
of the body except the germ cells (sperm and
egg) and therefore are not passed on to
children
Tumour suppressor gene
These genes normally function to PREVENT cell
growth/division
TS
____________________
____________________
____________________
____________________
____________________
Oncogene
Genes which normally function to PROMOTE cell
growth/division in a controlled manner
Ras
_________________
_________________
_________________
_________________
A nutritional example: the lactase gene
Many adult humans cannot metabolize
lactose (milk sugar). A single nucleotide
polymorphism (SNP), i.e., a one base-pair
difference in DNA sequence, correlates with
activation of the lactase promoter and with
lactose tolerance/intolerance.
agataatgtagTccctggcctca
agataatgtagCccctggcctca
ability to
activate
Oct-1
binding
phenotype
++
++
tolerant
+
+
intolerant
Olds, L. C. and E. Sibley (2003). Hum. Mol. Genet. 12(18): 2333-2340.
Now on to… Gene Regulation!
KEY CONCEPT
Gene expression is carefully regulated in both
prokaryotic and eukaryotic cells.
Control of Gene Expression
• Prokaryotic organisms regulate gene
expression ____________________________
• Eukaryotic cells regulate gene expression
_____________________________________
_
Control of Gene Expression
• Controlling gene expression is often
accomplished by controlling transcription
initiation
• Regulatory proteins bind to DNA to either
block or stimulate transcription, depending on
how they interact with RNA polymerase
Prokaryotic Regulation
• Prokaryotic cells often respond to their
environment by changes in gene expression
• Genes involved in the same metabolic
pathway are organized in _____________
• Some operons are _____________ when the
metabolic pathway is needed
• Some operons are _____________ when the
metabolic pathway is no longer needed
Prokaryotic cells turn genes on and off by
controlling transcription
• A promoter is a DNA segment that allows a gene to
be transcribed
• An operator is a part of DNA that turns a gene “on”
or ”off”
• An operon includes a promoter, an operator, and
one or more structural genes that code for all the
proteins needed to do a job
Prokaryotic Regulation
• Control of transcription initiation can be:
– ________________________– increases
transcription when ________________
bind DNA
– ________________________– reduces
transcription when ______________ bind
to DNA regulatory regions called
______________
lac Operon
– Operons are most common in
prokaryotes
– The lac operon was one of the first
examples of gene regulation to be
discovered
– The lac operon has three genes that code
for enzymes that break down lactose
lac Operon
• The lac operon contains genes for the use of
lactose as an energy source.
• Regulatory regions of the operon include the
CAP binding site, promoter, and the operator.
• The coding region contains genes for 3
enzymes
lac Operon
• The lac operon is negatively regulated by a
repressor protein:
– lac repressor binds to the operator to block
transcription
– in the presence of lactose,
_________________________________________
– repressor can no longer bind to operator
– transcription proceeds
lac Operon
• In the presence of both glucose and lactose,
bacterial cells prefer to use glucose
• Glucose prevents use of the lac operon
– binding of CAP – cAMP complex to the CAP binding
site is required for use of the lac operon
– high glucose levels cause low cAMP levels
– high glucose  low cAMP  lac operon _______
lac Operon
• The lac operon acts like a switch
– ___________________________________________
– ___________________________________________
trp Operon
• The trp operon encodes genes for the
biosynthesis of tryptophan
• The operon is not expressed when the cell
contains sufficient amounts of tryptophan
• The operon is expressed when levels of
tryptophan are low
trp Operon
• The trp operon is negatively regulated by the
______________________________
– trp repressor binds to the operator to block
transcription
– binding of repressor to the operator requires a
________________________ which is tryptophan
– low levels of tryptophan prevent the repressor
from binding to the operator
Eukaryotic Regulation
• Controlling the expression of eukaryotic genes
requires ____________________________
– general transcription factors are required for
transcription initiation
• required for proper binding of RNA polymerase to the
DNA
– specific transcription factors increase
transcription in certain cells or in response to
signals
Gene expression must be regulated in
several different dimensions—
In time:
10 wks
6 mos
14 wks
1 day
12 mos
18 mos
At different stages of the life cycle, different genes need to be on and off.
© M. Halfon, 2007
Eukaryotic Transcription
• General transcription factors bind to the
promoter region of the gene
• RNA polymerase II then binds to the promoter
to begin transcription at the start site
• Enhancers are DNA sequences to which
specific transcription factors (activators) bind
to increase the rate of transcription
• Transcription is controlled by regulatory DNA sequences
and protein transcription factors
– Most eukaryotes have a TATA box promoter
– Enhancers and silencers speed up or slow down the rate
of transcription
– Each gene has a unique combination of regulatory
sequences
Eukaryotic Chromosome Structure
• Methylation (the addition of –CH3) of DNA or
histone proteins
• Clusters of methylated cytosine nucleotides
bind to a protein that prevents activators from
binding to DNA
• Methylated histone proteins are associated
with inactive regions of chromatin
Protein Degradation
• Proteins are produced and degraded
continually in the cell
• Proteins to be degraded are tagged with
ubiquitin
• Degradation of proteins marked with ubiquitin
occurs at the proteasome